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中国农学通报

中国农学通报 ›› 2017, Vol. 33 ›› Issue (23): 6-9.doi: 10.11924/j.issn.1000-6850.casb17040113

所属专题： 园艺

• 生物技术科学 • 上一篇    下一篇

甜菜DAMD-PCR体系的建立及优化

兴旺,童乐怡,孙燕红,马龙彪,吴则东   

  1. 黑龙江省普通高校甜菜遗传育种重点实验室/黑龙江大学,黑龙江省普通高校甜菜遗传育种重点实验室/黑龙江大学,黑龙江省普通高校甜菜遗传育种重点实验室/黑龙江大学,黑龙江省普通高校甜菜遗传育种重点实验室/黑龙江大学,黑龙江省普通高校甜菜遗传育种重点实验室/黑龙江大学
  • 收稿日期:2017-04-18 修回日期:2017-06-08 接受日期:2017-06-23 出版日期:2017-08-21 发布日期:2017-08-21
  • 通讯作者: 吴则东
  • 基金资助:
    国家糖料现代农业产业技术体系甜菜高品质品种改良岗位专项基金“国家现代农业产业技术体系建设项目东北区育种岗位专家”(CARS- 210104)；农业部物种资源保护费项目“（甜菜种质资源收集鉴定编目繁种保存与利用）”(1120162130135252026)；科技部、财政部国家科技基础条件平 台项目“国家甜菜种质资源平台”(NICGR2016-017)。

Construction and Optimization of DAMD-PCR in Beet

  • Received:2017-04-18 Revised:2017-06-08 Accepted:2017-06-23 Online:2017-08-21 Published:2017-08-21

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336

摘要： 为了建立甜菜DAMD扩增体系，以期利用DAMD引物应用于甜菜品种指纹图谱的构建及分子标记辅助育种。本实验利用单因素变量的方法对甜菜DAMD体系进行优化。同时选用12 个甜菜品 种，利用优化的体系对25 条DAMD引物进行扩增。获得甜菜的最适DAMD体系：总体积为20 μL，包含模板DNA 10~80 ng、0.75 U 的DNA 聚合酶、0.2 μL 的dNTPs (2.5 mmol/L each)以及2.0 μL 的引物(10 μmol/L)。同时25 条引物均扩增出了清晰条带，除了个别引物多态性较差外，其余引物多态性都非常的丰富，其中引物62H(-)就可以把实验中用到的12 个甜菜品种全部区分开。由此可见，DAMD引物的扩增效率很高，并且扩增结果稳定，条带清晰，非常适合甜菜品种指纹图谱的构建及遗传多样性分析。

关键词: 铁皮石斛, 铁皮石斛, 腋芽, 原球茎, 激素

Abstract: To establish a beet DAMD amplification system and apply DAMD primer in construction of beet varieties fingerprint and molecular marker assisted breeding, this experiment optimized the beet DAMD system by using the single factor variable method. 12 beet varieties were chosen; the optimized system was used to amplify 25 DAMD primers. The optimum DAMD system of beet was as following: a total volume of 20 μL, template DNA 10-80 ng, 0.75 U of DNA polymerase, 0.2 μL of dNTPs (2.5 mmol/L each) and 2.0 μL of primers (10 μmol/L). At the same time, 25 primers were all amplified out of the clear bands, and except individual primers whose polymorphism was poorer, the other primers had very abundant polymorphism. The primers 62 H(-) could distinguish all 12 beet varieties. Thus, DAMD primers had high amplification efficiency, the amplification result was stable and the bands were clear, so DAMD primers were very suitable for the construction of fingerprint and the genetic diversity analysis of beet.